Continuous Administration of L-Dopa, Dopa Decarboxylase Inhibitors, Catechol-O-Methyl Transferase Inhibitors and Compositions for Same

ABSTRACT

Provided herein, in part, is a method of treating a neurological or movement disorder in a patient in need thereof, comprising subcutaneously administering to said patient a pharmaceutically acceptable composition comprising levodopa and optionally carbidopa and optionally entacapone or tolcapone, or pharmaceutically acceptable salts thereof, wherein said composition is administered substantially continuously, and compositions that can be used in the disclosed methods.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.16/749,435 filed Jan. 22, 2020, which is a continuation of U.S. patentapplication Ser. No. 15/209,423, filed Jul. 13, 2016, which is acontinuation of U.S. patent application Ser. No. 13/885,518, filed Oct.29, 2013 (now U.S. Pat. No. 9,421,267), which is the National Stagefiling under 35 U.S.C. § 371 of International Patent Application No.PCT/IL2011/000881, filed Nov. 15, 2011, which claims the benefit of andpriority to U.S. Provisional Application No. 61/413,637, filed Nov. 15,2010, and U.S. Provisional Application No. 61/524,064, filed Aug. 16,2011, each of which is hereby incorporated by reference in its entirety.

TECHNICAL FIELD

The present invention provides pharmaceutical compositions useful fortreatment of neurological or movement disorders such as Parkinson'sdisease, and a method for treatment such disorders by substantiallycontinuously subcutaneous administration of said compositions.

BACKGROUND

Parkinson's disease is a degenerative condition characterized by reducedconcentration of the neurotransmitter dopamine in the brain. Levodopa(L-dopa or L-3,4-dihydroxyphenylalanine) is an immediate metabolicprecursor of dopamine that, unlike dopamine, is able to cross theblood-brain barrier, and is most commonly used for restoring thedopamine concentration in the brain. For the past 40 years, levodopa hasremained the most effective therapy for the treatment of Parkinson'sdisease.

However, levodopa has a short half life in plasma that, even under bestcommon current standard of care, results in pulsatile dopaminergicstimulation. Long-term therapy is therefore complicated by motorfluctuations and dyskinesia that can represent a source of significantdisability for some patients. A therapeutic strategy that couldultimately deliver levodopa/dopamine to the brain in a more continuousand physiologic manner would provide the benefits of standard levodopawith reduced motor complications and is much needed by patientssuffering from Parkinson's disease and other neurological or movementdisorders (Olanow CW; Mov. Dis. 2008, 23(Suppl. 3):S613-S622).Sustained-release oral levodopa formulations have been developed, but,at best, such preparations have been found to be no more efficaciousthan standard tablets. Continuous administration of levodopa byintraduodenal administration or infusion has also been attempted byusing ambulatory pumps or patches. Such treatments, especiallyintraduodenal, are extremely invasive and inconvenient.

The metabolic transformation of levodopa to dopamine is catalyzed by thearomatic L-amino acid decarboxylase enzyme, a ubiquitous enzyme withparticularly high concentrations in the intestinal mucosa, liver, brainand brain capillaries. Due to the possibility of extracerebralmetabolism of levodopa, it is necessary to administer large doses oflevodopa leading to high extracerebral concentrations of dopamine thatcause nausea in some patients. Therefore, levodopa is usuallyadministered concurrently with oral administration of a dopadecarboxylase inhibitor, such as carbidopa or benserazide, which reducesby 60-80% the levodopa dose required for a clinical response, and thusprevents certain of its side effects by inhibiting the conversion oflevodopa to dopamine outside the brain. Various oral formulationstogether with inhibitors of enzymes associated with the metabolicdegradation of levodopa are well known, for example, decarboxylaseinhibitors such as carbidopa and benserazide, catechol-O-methyltransferase (COMT) inhibitors such as entacapone and tolcapone, andmonoamone oxidase (MAO)-A or MAO-B inhibitors such as moclobemide,rasagiline or selegiline or safinamide. Currently available oral drugsinclude SINEMET® and SINEMET® CR sustained-release tablets that includecarbidopa or levodopa; STALEVO® tablets containing carbidopa, entacaponeand levodopa; and MADOPAR® tablets containing levodopa and benserazide.There is an on-going and urgent need for methods and compositions thatcan effect continuous stimulation of L-dopa to more effectively treatmovement disorders such as Parkinson's disease. Nevertheless, no stableliquid formulation having e.g., an effective concentration in a volumesuitable for use for subcutaneous or transdermal delivery has ever beenachieved.

SUMMARY OF INVENTION

This disclosure generally relates, in part, to a pharmaceuticallyacceptable composition comprising 1) active components comprisingcarbidopa and at least about 4% by weight levodopa; and arginine andoptionally meglumine. Such compositions may have a pH of about 9.1 toabout 9.8 at 25° C.

In some embodiments, a disclosed composition having arginine may have amolar ratio of active components to the arginine is about 1:1.8 to about1:3.5, or about 1:2.3. In an exemplary embodiment, a disclosedcomposition may include about 4% to about 12% by weight or more oflevodopa and/or may include 1% to about 6% by weight carbidopa, e.g.about 1% to about 2% by weight carbidopa.

When meglumine is present in a disclosed composition, the molar ratio ofactive components to the arginine may be, for example, about 1:1.1 toabout 1:1.9, and the molar ratio of active components to the megluminemay about 1:0.3 to about 1:1.5, e.g., the molar ratio of activecomponents to the meglumine may about 1:0.3 to about 1:1.2, or forexample, about 1:0.4, or about 1:1.1. Such contemplated compositions mayinclude about 2.0% to about 11% by weight meglumine. Contemplatedcompositions as above for example may include 10% to about 35% by weightarginine.

Disclosed compositions may further comprise an agent that inhibits theformation of oxidation products, for example, such an agent may selectedfrom the group consisting of: ascorbic acid, Na-ascorbate, L-cysteine,N-acetylcysteine (NAC), gluthatione (GSH), Na₂-EDTA, Na₂-EDTA-Ca, andcombinations thereof. For example, the pharmaceutically acceptablecomposition disclosed herein may further include, in an exemplaryembodiment, ascorbic acid or a pharmaceutically acceptable salt thereof.In another or further embodiment, disclosed compositions may includesodium bisulfate.

Contemplated herein, for example, is a pharmaceutically acceptablecomposition comprising levodopa, arginine and optionally meglumine; andascorbic acid or a pharmaceutically acceptable salt thereof, e.g. thecomposition may have about 4% to about 12% by weight levodopa Theascorbic acid salt may selected, for example, from the group consistingof: ascorbate, sodium ascorbate, calcium ascorbate, potassium ascorbate,ascorbyl palmitate, and ascorbyl stearate. For example, a disclosedpharmaceutically acceptable composition may include the ascorbic acid ora pharmaceutically acceptable salt thereof is sodium ascorbate, e.g.,about 0.25% by weight or more ascorbic acid or a pharmaceuticallyacceptable salt thereof, about 0.2% to about 3% by weight ascorbic acidor a pharmaceutically acceptable salt thereof, or about 0.5% to about 1%by weight ascorbic acid or pharmaceutically acceptable salts thereof. Insome embodiments, a contemplated pharmaceutically acceptable compositionmay have a molar ratio of levodopa to the arginine is about 1:1.8 toabout 1:3.5, e.g., about 1:2.3.

Such contemplated compositions may further comprise carbidopa in someembodiments, for example, 1% to about 2% by weight carbidopa. In such anembodiment, the molar ratio of the levodopa and the carbidopa together,to the arginine, may be about 1:1.8 to about 1:3.5, e.g. about 1:2.3.Such a pharmaceutically acceptable composition may have a pH of about9.1 to about 9.8 at 25° C.

Contemplated compositions and formulations disclosed herein may be, forexample, liquid at room temperature. In some embodiments, a disclosedpharmaceutically acceptable composition may further comprise entacaponeor tolcapone.

Disclosed pharmaceutically acceptable formulations may be stable for atleast two weeks at 25° C.±5° C., and/or for example, may be stable forat least two months at −20° C.±5° C.

In an embodiment, a transdermal patch is contemplated herein suitablefor administering a disclosed pharmaceutically acceptable composition.

Also provided herein is a method of treating a neurological or movementdisorder in a patient in need thereof, e.g., Parkinson's disease,comprising administering to said patient a composition disclosed herein,e.g. a liquid composition of levodopa and/or carbidopa. Also providedherein, in one embodiment, is a method for treatment of a disease ordisorder such as a neurological disorder, or a disorder characterized byreduced levels of dopamine in a patient's brain, and/or for example adisorder such as Parkinson's disease, wherein the method includesadministration (e.g. substantially continuous administration) of adisclosed composition. In an embodiment, continuous administering mayinclude transdermal, intradermal, subcutaneous, intravenous,intrathecal, epidural, intracranial, or intraduodenal administration,e.g. may include the use of an infusion pump. Such methods may furthercomprise orally administering levodopa and/or carbidopa and optionallyentacapone or tolcapone.

Disclosed compositions may be administered subcutaneously and/or e.g.substantially continuously. Such subcutaneous administration maycomprise the use of one or more infusion pumps and/or transdermal and/ordermal patches. For example, a disclosed method may include a rate ofadministering a disclosed composition at least about 0.01 ml/hour toabout 0.2 ml/h, or at least about 0.07 ml/hour, or for example, about0.15 ml/hour during the day or during patient activity, and about 0 toabout 0.075 0.25 ml/hour at rest or sleep. Alternatively, a disclosedcomposition may be administered intraduodenally or intravenously.

In some embodiments, a method that includes subcutaneously administeringcomprises the use of one or more infusion pumps, e.g., with a rate ofadministering the composition is about 0.20 ml/hour to about 2.0 ml/h,for example, about 1.0±0.5 ml/hour., or about 1.25±0.5 ml/hour duringthe day or during patient activity, and about 0 to about 0.5 ml/hour atnight or at rest.

Also provided herein is a pharmaceutically acceptable compositioncomprising (i) carbidopa, at least 4% by weight levodopa, arginine andoptionally meglumine; or (ii) levodopa, arginine, optionally meglumine,and ascorbic acid or a pharmaceutically acceptable salt thereof, for usein treatment of a neurological or movement disorder. In one embodiment,the neurological or movement disorder is Parkinson's disease.

A pharmaceutically acceptable formulation is disclosed herein, in anembodiment, comprising about 2.5 to about 7% by weight levodopa, about 0to about 2% by weight carbidopa, about 5 to about 18% by weightarginine, and about 0.25% to about 3% by weight ascorbic acid or apharmaceutically acceptable salt thereof.

In an embodiment, a pharmaceutically acceptable formulation comprisingabout 8 to about 12% by weight levodopa, about 1 to about 3% by weightcarbidopa, about 15 to about 35% weight arginine is contemplated. Inanother embodiment, a pharmaceutically acceptable formulation comprisingabout 8 to about 12% by weight levodopa, about 1 to about 3% by weightcarbidopa, and about 12 to about 15% weight arginine, and about 3% toabout 10% by weight meglumine is provided. Such compositions may furtherinclude about 0.25-3% by weight ascorbic acid.

Also provided herein is a pharmaceutically acceptable liquid compositioncomprising arginine and at least about 7% by weight entacapone ortolcapone, e.g. at least about 8%, or at least about 10%, or about 7% toabout 12% by weight entacapone or tolcapone. For example, a disclosedcomposition may have entacapone or tolcapone and the arginine with amolar ratio of about 1:0.5 to about 1:2.5, for example about 1:1 toabout 1:1.5. Such liquid compositions may have a pH of about 6 to about9 at 25° C., and/or may be substantially stable at 25° C. for 48 hoursor more.

Provided herein, in an embodiment, is a process for preparing a stableliquid solution comprising levodopa and/or carbidopa, and arginine,comprising:

providing levodopa and/or carbidopa, and arginine to form a powdermixture;

adding water to said powder mixture to form a suspension;

heating said suspension at a temperature of about 40° C. to about 90° C.to form a solution; and

cooling said solution to provide the stable liquid composition. In someembodiments, wherein heating said suspension further comprises stirringthe suspension.

This disclosure relates at least in part to the discovery that argininecan form a salt of carbidopa, and/or levodopa and/or entacapone, ortolcapone, that can be used to form a stable, liquid formulation that issuitable for e.g., continuous subcutaneous, transdermal, intradermal,intravenous and/or intraduodenal administration. Such disclosedcompositions are capable of substantially continuously administeringcarbidopa, entacapone, tolcapone and/or levodopa to a patient in needthereof. For example, disclosed herein are compositions that relate tothe disclosed discovery that substantially continuously administering adopa decarboxylase inhibitor such as carbidopa, together withsubstantially continuously administering levodopa and optionallyentacapone or tolcapone, may stimulate L-dopa substantially continuouslyand thus e.g., extend the effectiveness of a levodopa oral dosingregimen and/or reduce the daily dosage of levodopa or eliminate the needfor oral levodopa, while effectively treating a movement and/orneurological disorder such as Parkinson's disease.

BRIEF DESCRIPTION OF THE FIGURES

FIGS. 1A and 1B depict the effect of carbidopa on the stability oflevodopa In Vitro and Ex Vivo: FIG. 1A. 6% weight levodopa and argininesolution with various concentrations (2, 1.5, 1, 0.5%) of carbidopa orno carbidopa were tested for physical stability in vitro. The resultsshow that carbidopa prevented dark yellow color formation in thepresence of air, in a dose related manner (small vials at the right handside), and in the absence of air (with N₂ in the head space) 0.5%carbidopa was sufficient to inhibit this color formation (large vials inthe left hand side of the figure). FIG. 1B. 7% weight percent levodopaand arginine solution, with or without 2% carbidopa by weight,continuously administered in to the subcutaneous tissue of a 5×5 cmfresh, full-thickness pig skin. The right hand side depicts theinhibition of oxidation with the use of a levodopa formulation thatincludes carbidopa.

FIG. 2 depicts that the presence of 1% carbidopa in a levodopa solutionreduces the severity and extent of local levodopa dependent subcutaneoustoxicity in the pig.

FIGS. 3A-3C depict the effect of carbidopa on the pharmacokinetics oflevodopa in the pig. FIG. 3A: the plasma concentration of levodopafollowing continuous subcutaneous administration of 6% levodopa withvarious amounts of carbidopa. FIG. 3B: The correlation between plasmasteady state concentration of levodopa, obtained following continuoussubcutaneous administration of levodopa/carbidopa formulations and theformulation concentration of carbidopa. FIG. 3C. The correlation betweenplasma steady state concentration of carbidopa following continuoussubcutaneous administration of levodopa/carbidopa formulations and theformulation concentration of carbidopa.

FIGS. 4A-4D show the effect of various agents on oxidation of levodopain the subcutaneous tissue of pig skin samples, ex-vivo, followingsubcutaneous administration of levodopa/carbidopa formulations.

FIG. 5A depicts the effect of continuous subcutaneous (SC) entacapone(200 mg/24 h) and/or carbidopa (CD) (40 mg/24 h) on the plasmaconcentrations of levodopa (ng/ml) following oral Administration ofSinemet (100/25 levodopa/carbidopa) in pigs.

FIG. 5B depicts the effect of continuous SC CD (40 mg/24 h) and/orlevodopa (LD) (140 mg/24 h) administration on the plasma concentrationsof levodopa following oral administration of Sinemet (100/25) in pigs.

FIG. 6 shows the effect of carbidopa on the local subcutaneous toxicityof levodopa following 24 h-continuous subcutaneous administration, at0.16 ml/h, in pigs.

DETAILED DESCRIPTION OF THE INVENTION

Disclosed herein, in an embodiment, is a liquid composition thatincludes an arginine salt of levodopa (e.g., arginine and levodopa), andoptionally carbidopa, that is stable at room temperature. Such disclosedcompositions may facilitate continuous delivery of an effective amountof levodopa, carbidopa, and/or other active agents such as entacapone ortolcapone to a patient in a minimally invasive fashion. Further,disclosed formulations have a pH that is suitable for e.g., transdermal,subcutaneous, intravenous, intrathecal, epidural, intracranial orintraduodenal administration.

For example, provided herein are formulations and methods capable ofobtaining substantially constant inhibition of COMT activity uponadministration, thereby increasing the half life of administeredlevodopa and substantially reducing the pulsatility of levodopa plasmalevels to avoid low trough levels of plasma levodopa.

Further, provided herein are formulations of levodopa and optionallycarbidopa that unexpectedly allow for stable dissolution of higherconcentrations (e.g., greater than 2% by weight) of levodopa at e.g. anacceptable pH, for e.g., substantially continuous subcutaneous ortransdermal administration. Such formulations may also be suitable forintravenous, intradermal, oral or intraduodenal administration. Forexample, provided herein are formulations and methods capable ofobtaining substantially constant plasma levodopa concentrations andsubstantially reducing the pulsatility of levodopa plasma levels toavoid low trough levels of plasma levodopa.

A treatment strategy of continuous levodopa and carbidopa (and/orentacapone or tolcapone) administration in accordance with the presentinvention may simulate L-dopa substantially continuously. For example,therapies and/or methods of the present invention may extend a levodopaoral dosing regimen to about 2 to about 3 times/day, and/or reduce dailydose of levodopa, and/or reduce or even eliminate oral dosing oflevodopa and carbidopa.

Compositions

Provided herein, in an embodiment, is a liquid formulation comprising anarginine salt of levodopa, or a liquid formulation comprising arginineand levodopa. In an embodiment, provided herein is a liquid formulationthat includes levodopa and arginine in a molar ratio of about 1:1.5 toabout 1:2.5, or about 1:2 to about 1:2.3 levodopa:arginine, or, forexample, when such a liquid composition further comprises carbidopa in amolar ratio of about 1:2 to about 1.3.5, or about 1:1.8 to about 1:3.5carbidopa:arginine.

Such levodopa and arginine formulations or solutions may have a pH thatis pharmaceutically acceptable for subcutaneous administration, e.g. apH of about 8 to about 10, for example, about 9.1 to about 9.8, e.g.,9.2 to 9.6 at 25° C. A disclosed formulation having levodopa andarginine may include at least about 7%, 8%, 9%, or more by weightlevodopa, e.g., may include about 10%, 20% or more by weight levodopa.In some embodiments, a disclosed formulation may include about 2.5 toabout 10 weight percent levodopa, 4 to about 7 weight percent levodopa,or about 7.5 to about 12 weight percent levodopa, or about 5% to about30%, or about 10 to about 20 weight percent levodopa, and may furtherinclude about 9 to about 20 weight percent arginine or about 9 to about30 weight percent arginine, e.g. about 10 to about 18 weight percentarginine, about 10 to about 20% or about 15 to about 30% or more byweight arginine or about 12, 13, 14, or 15 weight percent arginine. Forexample, arginine may be present in contemplated formulations at a molarratio of about 1.5:1 to about 3:1, e.g. 1.8:1 to about 3.5:1, ratio ofarginine:total active ingredients (which may include e.g., levodopa,carbidopa, etc.).

For example, disclosed herein is a pharmaceutically acceptablecomposition, having a pH of about 9.1 to about 9.8 at 25° C., thatincludes the active components levodopa and carbidopa (e.g. about 4% byweight or more levodopa), and arginine and/or meglumine. For example,contemplated compositions having levodopa and arginine may furthercomprise carbidopa, for example, may further include at least about 1%,at least about 2%, at least about 4% by weight carbidopa, for exampleabout 2% to about 4% by weight carbidopa. For example, provided hereinis a composition comprising arginine and about 2% to about 12% by weightlevodopa or more (e.g. about 4% to about 10%, about 4% to about 7%,about 5% to about 10%, or about 6% to about 11% by weight levodopa, orabout 5% to about 20% by weight levodopa) and about 1% to about 6%,about 1% to about 2% (e.g. about 1.25 or about 1.5%), or about 2% toabout 5% or about 2% to about 4% by weight carbidopa. When administeredsubcutaneously and/or dermally, such compositions having levodopa andcarbidopa may result in minimal local tissue damage, e.g., as comparedto subcutaneous or dermal administration of a composition that includeslevodopa (e.g., a levodopa/arginine composition) alone. Further, suchlevodopa and arginine compositions, when further including carbidopa,may have more stability (e.g. may not form unwanted oxidation productsover time as compared to a composition having levodopa and argininealone).

In another embodiment, disclosed formulations may include an amino sugarsuch as meglumine, which may, for example, replace some or all of thearginine present in the formulations. For example, disclosed here is aformulation comprising levodopa and/or carbidopa and meglumine. Alsocontemplated herein is a meglumine salt of levodopa and a meglumine saltof carbidopa. In an embodiment, a composition comprising arginine andhaving active agents such as levodopa and carbidopa, wherein the molarratio of active agents to arginine is less than about 1:2; to improvethe stability of such compositions, this exemplary composition mayfurther comprise meglumine, e.g., with a ratio of active agents tomeglumine of about 1:0.3 to about 1.1.5. For example, provided herein isa composition having levodopa or carbidopa (or a combination) as activecomponents, arginine, and meglumine, wherein the molar ratio of activecomponents to arginine is about 1:1.1 to about 1:1.9 (e.g. 1:1.3) andthe molar ratio of active components to meglumine of about 1:0.3 toabout 1:1.2 (e.g. about 1:0.4, 1:0.5, 1:0.8, 1:1.1). Contemplatedcompositions can include levodopa (e.g. about 4 to about 10% by weightor more), carbidopa (e.g. about 0.5 to about 3% by weight, e.g. about 1or 2% by weight), about 9% to about 16 by weight arginine, and about 2%to about 10% by weight meglumine.

Also provided herein, in an embodiment, is a formulation comprisinglevodopa, arginine, and/or carbidopa, and optionally for example anagent that inhibits the formation of oxidation products. Such aformulation may be liquid at room temperature, with a pH of about 9.1 to9.8. For example, provided herein is a composition that includesascorbic acid or salt thereof.

In an embodiment, a disclosed composition may further comprise one ormore agents that inhibit the formation of oxidation products. Such agentmay be e.g., tyrosinase inhibitors and/or o-quinone scavengers and/orCu⁺⁺ chelators and/or antioxidants. In some embodiments, carbidopa mayact as an agent that inhibits the formation of oxidation products. Forexample, contemplated formulations may include o-quinone scavengers suchas, but not limited to, N-acetyl cysteine, gluthatione, ascorbic acid,Na-ascorbate, and/or L-cysteine. In an embodiment, formulations mayinclude an agent chosen from one or more of tyrosinase inhibitors suchas captopril; methimazole, quercetin, arbutin, aloesin,N-acetylglucoseamine, retinoic acid, α-tocopheryl ferulate, MAP (Mgascorbyl phosphate), substrate analogues (e.g., sodium benzoate,L-phenylalanine), Cu′ chelators for example, Na₂-EDTA, Na₂-EDTA-Ca, DMSA(succimer), DPA (D-penicillamine), trientine-HCl, dimercaprol,clioquinol, sodium thiosulfate, TETA, TEPA, curcumin, neocuproine,tannin, and/or cuprizone. Other contemplated anti-oxidants that may formpart of a disclosed formulation include sulfite salts (e.g., sodiumhydrogen sulfite or sodium metabisulfite), di-tert-butyl methyl phenols,tert-butyl-methoxyphenols, polyphenols, tocopherols and/or ubiquinones,including but not limited to caffeic acid.

In a particular embodiment, provided herein are compositions thatinclude levodopa, carbidopa, arginine, optionally meglumine, and anascorbic acid or pharmaceutically acceptable salt thereof. For example,contemplated compositions may further include ascorbate, sodiumascorbate, potassium ascorbate, calcium ascorbate, ascorbyl stearate,and/or ascorbyl palmitate. For example, a composition may include about0.5 percent by weight or more (e.g., about 0.5 to about 3 percent byweight, or about 0.2 to about 2 percent or about 0.5 to about 1 percentby weight, e.g. about 0.75% by weight ascorbic acid or salt thereof.

Provided herein, in an embodiment, is a pharmaceutically acceptableformulation that includes entacapone (or tolcapone), and arginine, thatallows for substantially continuous administration of entacapone ortolcapone. For example, provided herein, for example, is a stable liquidformulation that includes entacapone or tolcapone and may be suitablefor substantially continuous administration to a patient. Further, suchformulations may have a physiologically acceptable pH, for example,about 6 to about 9.5, or about 6.5 to about 8.5, or about 7 to about 8.

For example, entacapone (or tolcapone) and arginine may be dissolved inan aqueous solution, (e.g., having a pH of about 6 to 9, e.g., fromabout 6.5 to about 8.5, e.g., from about 7 to 8 at 25° C. or at 30° C.Alternatively, entacapone (free base) (or tolcapone (free base) and abasic amino acid salt (e.g. arginine and/or lysine) are dissolvedtogether in a liquid (e.g. an aqueous liquid) to form a disclosed liquidformulation. Disclosed liquid formulations may include about 2% byweight entacapone or tolcapone, about 4% by weight entacapone ortolcapone, or about 2% to about 12% by weight entacapone or tolcapone,for example, may include about 7% by weight or more, about 8% by weightor more, or about 10% by weight or more entacapone or tolcapone, forexample, may include about 3% to about 20% by weight or more entacaponeor tolcapone, e.g., about 5% to about 8% by weight, about 8% to about12% by weight entacapone or tolcapone. For example, a liquid formulationmay include entacapone, and a basic amino acid (such as arginine) inmolar ratio of about 1:0.5 to about 1:2.5, or about 1:1 to about a 1:2,e.g., about 1:1 or 1:1.5. Such liquid formulations may further comprisecarbidopa, for example, at least about 2% by weight or at least about 4%by weight carbidopa, e.g. about 2% to about 6% or more by weightcarbidopa. In another embodiment, such liquid formulations may furthercomprise levodopa, for example, at least about 2%, 3%, 4%, 5%, 6%, or 7%by weight levodopa, e.g. about 2.5% to about 12% by weight levodopa. Inan exemplary embodiment, a composition that includes tolcapone orentacapone may further include an excipient such as an α, β orγcyclodextrin or derivative.

Disclosed liquid formulations (e.g. a liquid composition comprisinglevodopa, carbidopa, entacapone tolcapone, or combinations of two ormore) and arginine (and/or meglumine), e.g., a disclosed formulationcomprising levodopa and arginine) may be stable for 24 hours, for 48hours, for 7 days, or more at 25° C. For example, an exemplary liquidformulation may include about a 1:1 molar ratio of entacapone:arginine(or tolcapone:arginine), with about 5% to about 15%, or about 6% toabout 12%, or 6% to about 10% by weight entacapone. Such an entacapone,arginine liquid formulation may be more stable, in some embodiments, at7 days as compared to a liquid composition that includes a lysine orhistidine salt of entacapone. In an embodiment, a disclosed formulationcomprising levodopa and arginine may be stable for at least one week, orat least two weeks or more at room temperature, e.g. at 20° C. to 30°C., e.g. at 25° C. In an embodiment, a disclosed formulation comprisinglevodopa and arginine may be stable for at least one month, or at leasttwo months at temperature below freezing e.g. at −10° C. and/or at −20°C., at −18° C., or e.g., at −20 to −80° C. The term “stable” in thiscontext means that a formulation does not significantly precipitate outof solution and/or one or more active agents does not degradesignificantly for a substantial amount of time.

In some embodiments, disclosed liquid formulations or compositions areliquid solutions, i.e. are substantially homogenous liquid mixtures.Such liquid mixtures may comprise water and/or other pharmaceuticallyacceptable excipients. In another embodiment, disclosed liquidcompositions may be substantially non-aqueous.

In some embodiments, a disclosed liquid formulation will be stable for aperiod of 1 day, 2 days, 3 days, 1 week, or 1 month or more at roomtemperature. In an embodiment of the invention, a disclosed liquidformulation further comprise a pharmaceutically acceptable excipientsuch as e.g., N-methylpyrrolidone (NMP), or polyvinylpyrrolidone (PVP),EDTA (or salts thereof) cysteine, N-acetylcysteine and/or sodiumbisulfate.

For example, in one embodiment, provided herein is a stable liquidformulation that comprises about 4% to about 12% by weight levodopa,and/or carbidopa (e.g. about 1% to about 6% by weight, or about 2% toabout 6% by weight) and/or entacapone or tolcapone (e.g. about 7% toabout 12% by weight) and about 1 to about 40% arginine, about 0 to about10% NMP, about 0 to about 5% PVP, and/or about 0 to about 3.5% of one ormore water soluble antioxidants, by weight.

The invention further provides a stable lyophilized powder comprising anarginine salt of levodopa, carbidopa or entacapone, or a combination oftwo or more of levodopa, carbidopa or entacapone. In one embodiment,such stable lyophilized powder may comprise about 20-99% of the levodopaor entacapone salt, about 0-60% NMP, about 0-15% PVP, and about 0-10% ofone or more water soluble anti oxidants. The lyophilized powder can bereconstituted into a liquid formulation by addition of water alone orwater with NMP, and may include or not include antioxidants.

In some embodiments, provided herein is a formulation suitable forcontinuous subcutaneous administration, e.g., comprising about 4 toabout 7% by weight levodopa, about 1 to about 2% by weight carbidopa,and about 10 to about 18% weight percent arginine. Such formulations mayfurther include, in some embodiments, about 1% weight percent ascorbicacid (or a pharmaceutically acceptable salt thereof), and/or optionallyabout 0.2% weight percent Na₂EDTA-Ca.

In another embodiment, a formulation suitable for intraduodenal orintravenous administration is provided comprising at least about 8% byweight levodopa (e.g. 8% to about 12%), at least about 1.5% by weightcarbidopa (e.g. about 1.5% to about 3%), and at least about 15% arginineby weight (e.g. about 15% to about 30% by weight, or about 15% to about20% by weight). Such formulations may include about 1% by weightascorbic acid and about 0.2% by weight Na₂EDTA-Ca. Also provided hereinis a formulation suitable for intraduodenal or intravenousadministration, comprising at least about 8% by weight levodopa (e.g. 8%to about 12%), at least about 1.5% by weight carbidopa (e.g. about 1.5%to about 3%), at least about 10% arginine or about 12% arginine byweight (e.g. about 10% to about 15% by weight, or about 12% to about 15%by weight), at least about 3% by weight meglumine (e.g. about 3% toabout 8%, or about 3% to about 5%), and optionally about 1% by weightascorbic acid (or salt therefore), and/or optionally about 0.2%Na₂EDTA-Ca for continuous intraduodenal or intravenous administration.Such formulations may include water.

Liquid formulations of the invention may be designed for continuousadministration of entacapone, tolcapone, carbidopa and/or levodopa apatient in need thereof. For example, a patient may be substantiallycontinuously administered (e.g. subcutaneously, transdermally,intraduodenally, intradermally, or intravenously) a formulation thatincludes a disclosed entacapone composition that includes arginine andentacapone, while carbidopa, a carbidopa salt, or a compositioncomprising carbidopa is also substantially continuously administerede.g. a different device, or in a separate compartment in the samedevice, in a separate composition via the same device, or in the samecomposition, and/or optionally levodopa and/or carbidopa is orallyadministered at discrete intervals, e.g., 2, 3, 4, or 5 times a day.

As used herein in the specification, the term “a composition comprisinglevodopa” or “levodopa composition” contemplates formulations thatcomprise levodopa, optionally together with a decarboxylase inhibitor, acatechol-O-methyl transferase (COMT) inhibitor, and/or a MAO-A or MAO-Binhibitor. For example, a composition comprising levodopa includes adosage formulation that comprises levodopa (or a salt thereof) andoptionally another drug, where the dosage formulation may be animmediate release, controlled release, dual release or multiple releaseformulation suitable for oral administration.

The term “decarboxylase inhibitor” refers to a dopa decarboxylaseinhibitor, e.g., a drug that inhibits the peripheral metabolism oflevodopa to dopamine by aromatic L-amino acid decarboxylase such ascarbidopa and benserazide.

A movement disorder refers to a nervous system condition that causesabnormal voluntary or involuntary movements, or slow, reduced movements.

A neurological disorder is a disorder of the body's nervous system.

The term “pharmaceutically acceptable carrier” or “pharmaceuticallyacceptable excipient” as used herein refers to any and all solvents,dispersion media, preservatives, antioxidants, coatings, isotonic andabsorption delaying agents, and the like, that are compatible withpharmaceutical administration. The use of such media and agents forpharmaceutically active substances is well known in the art. Thecompositions may also contain other active compounds providingsupplemental, additional, or enhanced therapeutic functions.

Pharmaceutically or pharmacologically acceptable” include molecularentities and compositions that do not produce an adverse, allergic orother untoward reaction when administered to an animal, or a human, asappropriate. For human administration, preparations should meetsterility, pyrogenicity, general safety and purity standards as requiredby e.g, the U.S. FDA Office of Biologics standards.

The term “pharmaceutical composition” as used herein refers to acomposition comprising at least one active agent as disclosed hereinformulated together with one or more pharmaceutically acceptablecarriers.

The term “physiologically acceptable pH” is understood to mean a pH ofe.g., a composition that facilitates administration of the compositionto a patient without significant adverse effects, e.g. a pH of about 4to about 9.8, (for example, about 4±0.3 to about 9.5±0.3.)

COMT inhibitors refer to inhibitors that inhibit the degradation oflevodopa to 3-methyldopa by catechol-O-methyl transferase and prolongthe action of levodopa, such as such as entacapone or tolcapone. Forexample, compositions comprising levodopa contemplated herein may alsoinclude a decarboxylase inhibitor (carbidopa or benserazide) andentacapone, e.g. “triple therapy”.

MAO-A or MAO-B inhibitors prevent the breakdown of dopamine by monoamineoxidases, e.g., moclobemide, rasagiline, selegiline or safinamide, e.g.,rasagiline.

Kits and Devices

Contemplated herein, in part, is a transdermal patch suitable fortransdermal or subcutaneous administration of an active agent thatcomprises a composition as disclosed herein, for example, may include acomposition including levodopa and carbidopa, and arginine, as disclosedherein, and optionally a composition that comprises carbidopa and/orlevodopa. Such patches may have one or more compartments which may havethe same or different compositions, for example, one compartment mayhave a disclosed formulation and another a different disclosedformulation, or a different active formulation. A transdermal patchrefers to any device that is capable of delivering one or more of theactive agents forming a disclosed composition through the skin or mucousmembrane into the bloodstream of a patient.

Also contemplated herein is a kit comprising: a) a first formulationcomprising a disclosed composition comprising carbidopa and arginine,wherein said first formulation is suitable for continuous (e.g dermal orsubcutaneous) administration; optionally b) a second formulationcomprising levodopa or an arginine salt of levodopa, wherein the secondformulation is suitable for continuous administration; optionally c) athird formulation comprising entacapone and arginine, wherein the thirdformulation is suitable for continuous administration, and/or optionallyd) a fourth formulation comprising tolcapone and arginine, wherein thefourth formulation is suitable for continuous administration, optionallye) a fifth composition comprising levodopa and optionally carbidopa,wherein said fifth formulation is suitable for subcutaneousadministration; and/or optionally f) a sixth composition comprisinglevodopa and/or optionally carbidopa, wherein said sixth formulation issuitable for oral administration; and g) instructions for administrationof at least one of formulations a)-f). The formulations a)-e) may besuitable for continuous administration by any suitable route such astransdermally, intravenously, subcutaneously, intradermally,intramuscularly or intraduodenally.

In an embodiment, the first formulation comprises a disclosed carbidopasalt and is suitable for administration subcutaneously. The sixthformulation of a contemplated kit may include levodopa, a levodopa salt,or a composition comprising levodopa, and may be presented as anysuitable oral dosage such as, but not limited to, pills, tablets,dispersible tablets, capsules, liquid, and the like. In an embodiment,the fourth formulation may be in the form of an immediate release,controlled release or dual release oral formulation that comprises bothlevodopa and benserazide, or both levodopa and carbidopa. Such oralformulation in the form of pills, tablets, or the like, may comprise aratio of carbidopa or benserazide to levodopa of about 1:10 to 1:4,preferably from about 1:4 to 1:1. Other contemplated second formulationsinclude formulations, e.g., tablets that include levodopa, carbidopa,and entacapone (or tolcapone), or e.g. a tablet that includes levodopaarginine salt and/or carbidopa arginine.

A contemplated kit may include a levodopa arginine salt (and/orcarbidopa arginine salt), or a liquid composition having levodopa,carbidopa, and/or entacapone (or a combination) and arginine. Suchcomposition may be liquid or a lyophilized powder that can bereconstituted into a liquid formulation, or, for example, may form partof a transdermal patch, and may be designed for continuousadministration by any suitable route such as, but not limited to,transdermally, intravenously, subcutaneously, intradermally,intramuscularly or intraduodenally.

In another embodiment, the kit comprises a first liquid formulationcomprising carbidopa and arginine (and optionally levodopa and/orentacapone or tolcapone) suitable for, but not limited to, transdermal,intravenous, subcutaneous, intradermal, intramuscular, intraduodenalcontinuous administration, and a second formulation in the form of animmediate release, controlled release or dual release oral formulationcomprising levodopa and carbidopa and/or a second liquid formulationcomprising entacapone and arginine (or tolcapone and arginine), suitablefor, but not limited to, transdermal, intravenous, subcutaneous,intradermal, intramuscular, intraduodenal continuous administration.

In some embodiments, disclosed liquid compositions (e.g. comprisinglevodopa, arginine and optionally carbidopa), may be provided in e.g. apre-filled cartridge suitable for use by a patient or physician. Forexample, provided herein is a kit comprising a prefilled cartridgewherein a disclosed liquid formulation is disposed within the cartridge(e.g., a pre-filled cartridge having a single dose or a dose suitablefor a single administration to a patient of a levodopa and argininesolution (and optionally carbidopa)), and optionally instructions foruse.

Preparation of Compositions

Disclosed formulations or compositions may be prepared by mixingarginine and/or meglumine in amounts as disclosed above with levodopaand/or carbidopa, and optionally anti oxidant(s) e.g., to form a powdermixture. Water may be added to the mixture to form a suspension. Thesuspension may be heated to about e.g., at about 40 to about 100° C., orat about 60 to 90° C., e.g., 72±5° C.) e.g., by adding pre-heated waterand/or by placing the mixture in a hot (e.g. 72±5° C.) water bath (e.g.for about 3, about 5, about 10 minutes or more (e.g. up to about 10minutes)), to form a solution, with optional stirring, and cooling thesolution to form the composition. N₂ may be provided the head space ofthe container. For example, the mixture can then be removed from the hotwater bath, and cooled to room temperature, and adding, e.g.,immediately thereafter, an optional anti-oxidant(s) under N₂ atmosphereand subsequent stirring. A preparation such as that above, e.g., wherelevodopa, carbidopa, and arginine are mixed together as powders first,and a suspension formed with water and then heated may result in a morestable solution as compared to a preparation that includes a step wisepreparation of individual water suspensions of ingredients and latercombination.

Disclosed formulations can be sterilized, e.g., using 0.2 μM filterssuch as filters with nylon or PVDF membranes. In some embodiments, thepreparation of disclosed formulations has fewer undesirable by-products(e.g. toxic by-products) or contaminants (for example, hydrazine) whencarbidopa and levodopa are present at the same time and/or when preparedusing certain antioxidants (e.g. ascorbic acid or salts thereof) ratherthan others (e.g. sodium bisulfite). In another embodiment, thepreparation of disclosed formulations has fewer undesirable by-productswhen pre-heated water is added as disclosed above, as compared to aformulation prepared without the addition of pre-heated water. Inanother embodiment, the levodopa and/or carbidopa may not dissolveunless the preparation procedure disclosed is used. Such disclosedpreparations as above may provide a more stable formulation as comparedto a formulation prepared without adding hot water or heating.

Methods of Treatment

In a further aspect, the present invention provides a method fortreatment of a disease or disorder, such as a neurological or movementdisorder, comprising substantially continuously administering adisclosed composition, and/or administering composition comprisinglevodopa (e.g. orally administering a levodopa composition orsubcutaneously administering such as a disclosed levodopa composition),and optionally co-administering substantially continuously to a patientin need a therapeutically effective amount of a composition comprising adecarboxylase inhibitor or a salt thereof (e.g. comprising carbidopa andarginine), optionally together with substantially continuouslyadministering a composition such as disclosed herein, comprising atherapeutically effective amount of a COMT inhibitor (e.g a entacaponecomposition comprising entacapone and arginine, or a tolcaponecomposition comprising tolcapone and arginine). In some embodiments, aprovided method may comprise substantially continuously administering acomposition that includes both e.g. carbidopa and entacapone ortolcapone, or may comprise substantially continuously administering twoseparate compositions (e.g., one having entacapone or tolcapone, onehaving carbidopa and/or levodopa), such as the compositions disclosedherein. As shown in the Examples, patients administered levodopatogether with continuous administration of carbidopa and entacapone mayresult in higher plasma levels of levodopa as compared to the plasmalevels a person of skill in art would expect from continuousadministration of carbidopa or entacapone alone.

For example, provided herein are methods of treating neurological ormovement disorders that include oral administration of a compositioncomprising levodopa and/or carbidopa, (and optionally, oraladministration of a composition comprising a COMT inhibitor), and alsoincludes subcutaneous administration of a carbidopa composition such asdisclosed herein, or composition that includes both levodopa and/orcarbidopa, such as disclosed herein (and where subcutaneousadministration of levodopa and carbidopa may be as separate compositionsor compositions that include both levodopa and carbidopa). Such methodscan also include subcutaneous administration of levodopa and/or a COMTinhibitor, which may be in different formulations or in the formulation.

Also provided herein are methods of treating neurological or movementdisorders that include subcutaneous or dermal, substantially continuousadministration of a composition (e.g. a liquid composition) comprisinglevodopa and optionally carbidopa (and may include optionallysubstantially administering a composition comprising carbidopa), evenwithout discrete (e.g. oral) administration of levodopa, which may besufficient to maintain therapeutic plasma levels of levodopa. In anembodiment, contemplated methods may include substantially continuouslyadministering levodopa and carbidopa, (in the same composition orseparate compositions), e.g. with levodopa:carbidopa weight ratios ofabout 10:1 to about 1:1. In an embodiment, contemplated methods mayinclude substantially continuously administering levodopa with orwithout carbidopa and/or oral COMT inhibitors.

In some embodiments, compositions comprising levodopa (e.g. a disclosedliquid composition) may be administering at a rate of about 0.07ml/hour, or e.g., about 0.01 ml/hour to about 0.2 ml/hour. Such ratesmay be constant throughout the day and night or varied according topatient's need, for example, may reflect a patient resting or sleepingschedule and waking or higher activity level schedule. For example,liquid compositions such as those disclosed herein (e.g includinglevodopa) may be administered at a rate of about 0.15 ml/hour in themorning (e.g. for about 2-3 hours before waking), about 0.1 ml/hoursduring the daytime or activity time, (e.g. for about 10 to about 12hours), and/or about 0.035 ml/hour at rest or at night. In anotherembodiment, liquid composition such as those disclosed herein (e.g.,disclosed compositions comprising levodopa) may be administered, e.g.,intraduodenally, at a rate of about 1.0 ml/hour during the daytime oractivity time (e.g. for about 2-3 hours before waking and for about 10to about 12 hours thereafter), and 0 to about 0.5 ml/hour at rest or atnight. In another embodiment, liquid compositions such as disclosedherein (e.g. comprising levodopa and arginine), may be administered at arate of about 1.25 ml/hour (e.g. about 1.25±0.5 ml/hour during thedaytime or activity time (e.g. for about 2-3 hours before or afterwaking and for about 10 to about 14 hours thereafter) and 0 to about 0.5ml/hour (e.g. about 0.5±0.25 ml/hour) at rest or night.

Contemplated administration of e.g., carbidopa, entacapone, tolcapone,and/or levodopa, following the disclosed methods, typically can becarried out over a defined time period (usually weeks, months or yearsdepending upon the combination selected). Contemplated therapies areintended in part to embrace administration of multiple therapeuticagents in a manner wherein a dopa decarboxylase inhibitor and optionallya COMT inhibitor (e.g. entacapone or tolcapone) is administeredsubstantially continuously while levodopa is administered at discreteintervals, as well as administration of contemplated therapeutic agents,or at least two of the therapeutic agents (e.g. levodopa and carbidopa,and optionally entacapone or tolcapone, or levodopa and entacapone ortolcapone) in a substantially simultaneous manner, which may beadministered in the same composition or e.g., simultaneouslyadministered but as different compositions. Administration can beeffected by any appropriate route including, but not limited to, oralroutes, intravenous routes, intramuscular routes, intradermal routes,subcutaneously, transdermally, and direct absorption through mucousmembrane tissues.

In some embodiments, levodopa can be administered by the same route orby different routes as compared to administration of e.g. a contemplatedcarbidopa formulation. For example, carbidopa may be administeredsubcutaneously, e.g., substantially continuously, while levodopa may beadministered orally, e.g. at discrete intervals. In an embodiment, adisclosed liquid carbidopa composition (e.g. having carbidopa andarginine) and a liquid entacapone composition (e.g. having entacaponeand arginine) is administered substantially continuously, while an oralcomposition that includes levodopa (and may also include one or moreother active agents such as a dopa decarboxylase inhibitor and/or a COMTinhibitor) is administered at discrete intervals. Alternatively, forexample, both levodopa and carbidopa may be administered subcutaneouslyor transdermally. Disclosed compositions may be administeredsubstantially continuously over 12 hours, 1 day, 1 week, or more.

The disease or disorder characterized by reduced levels of dopamine inthe brain contemplated herein are neurological or movement disordersincluding restless leg syndrome, Parkinson's disease, secondaryparkinsonism, Huntington's disease, Shy-Drager syndrome and conditionsresulting from brain injury including carbon monoxide or manganeseintoxication. Methods for treating such disorders in a patient in needthereof are provided, for example, by administering (e.g.,subcutaneously) a disclosed composition. In one embodiment, the diseaseto be treated is Parkinson's disease.

In an embodiment, substantially continuously administering using e.g. aliquid formulation may be via a pump for subcutaneous infusion (insulinpump) at an average rate of about 10-250 μl/hour, or about 160±40μl/hour continuously for 24 hours; about 200±50 μl/hour continuously for16 hours (during waking hours) and at night (e.g. for 8 hours, about 0to 80 μl/hour or via a transdermal patch. In an embodiment,substantially continuously administering intravenously orintraperitonealy using e.g. a liquid formulation may be at an averagerate of about 0.2-2 ml/hour, or about 1±0.5 ml/hour continuously for 24hours; about 1.0±0.5 ml/hour continuously for 16 hours (during wakinghours) and at night (e.g. for 8 hours, about 0 to 0.5 ml/hour via a pumpor transdermal patch, or combination of delivery devices that aresuitable for e.g. subcutaneous, intravenous, intrathecal, or via theduodenum).

The invention now being generally described, it will be more readilyunderstood by reference to the following examples which are includedmerely for purposes of illustration of certain aspects and embodimentsof the present invention, and are not intended to limit the invention inany way.

EXAMPLES Example 1: Preparation of Solutions/Formulation forSubcutaneous Administration

A. A 2% Carbidopa solution/formulation was prepared by adding pre-heated0.1% Na-bisulfite solution to carbidopa [ASSIA Ltd.]. Arginine [MERCK]was added to obtain a final molar ratio of 1:1.2 CD(carbidopa):arg(argininine). The mixture was stirred at 60° C. untilcomplete dissolution was obtained. Heating was stopped and thepreparation was allowed to cool down to room temperature pH of 8.5.Solution was filtered using a sterile 0.2204 PVDF membrane.

B. A 10% tolcapone solution/formulation was prepared as follows: asolution containing 10% tolcapone was prepared by adding the respectiveamount of H₂O to tolcapone [Synfine Research], slowly adding argininewhile stirring to obtain a final molar ratio of 1:1. The mixture isstirred until complete dissolution is obtained. After cooling down, thepH of the solution was 7.8.

C. A solution containing 10% entacapone was prepared by adding therespective amount of H₂O to entacapone [Suven Life Sciences], stirringat 30-35° C. and slowly adding arginine to obtain a final molar ratio of1:1. The mixture is stirred until complete dissolution is obtained.After cooling down, the pH of the solution was 6.9. The pH of lessconcentrated solutions (6%) was 7.8. After preparation, such entacaponesolution can be diluted to a 2%, 3% or 4% by weight formulation.

Entacapone did not dissolve (at concentrations >1%) with other aminoacids such as histidine and glutamic acid or in buffers at various pHs.

D. A 7% levodopa/2% carbidopa solution was prepared by adding pre-heated0.1% Na-bisulfite solution to arginine. Levodopa was added to obtain afinal molar ratio of 1:2 LD:arg. The mixture was stirred at 75-80° C.until complete dissolution was obtained. After cooling down to 60° C.,carbidopa and arginine were added to obtain a final molar ratio of 1:1.2CD(carbidopa):arg(arginine). The mixture was stirred at 60° C. untilcomplete dissolution was obtained. After cooling, about 12.5% morearginine was added to the solution. The pH of the solution was about9.2.

E. A 7% weight percent. Levodopa solution was prepared by addingpre-heated 0.1% Na-bisulfite solution to arginine. Levodopa was added toobtain a final molar ratio of 1:2 LD:arg. The mixture was stirred at75-80° C. until complete dissolution was obtained. After cooling down,the pH of the solution was about 9.4.

Example 2: Formulation Preparation Procedure

Levodopa (LD) and carbidopa (CD) formulations can be prepared asfollows. However, as shown in Table A, the method of preparation hassignificant impact on the resulting composition's physical and chemicalstability.

Method #1(L-Arg solution): L-Arg and Na-Bis (Na-bisulfate) weredissolved in water. The solution was added to the LD and CD powders. Themixture was heated with stirring for 13 min at 75° C. until fullydissolved. LD/CD solution kept at room temperature (RT) for 10 min tocool down.

Method #2 (all powders together): All powders (LD, CD and L-Arg) wereweighed and water with Na-Bis was added. Suspension was heated withstirring for 13 min at 75° C. until fully dissolved. LD/CD solution keptat RT for 10 min to cool down.

Method #3 (same as #2 without Na-Bis pre-heating): All powders (LD, CDand L-Arg) were weighed together and water was added. Suspension washeated with stirring for 13 min at 75° C. until fully dissolved. LD/CDsolution kept at RT for 10 min to cool down.

Method #4 (preparation in steps): LD and the respective amount of L-Argwere weighed; water and Na-Bis solution were added. The suspension washeated for 7 min at 75° C. until fully dissolved followed by 7 min atRT. CD and the respective amount of L-Arg were weighed, and added to theLD-arg solution at 60° C. until fully dissolved. Finally, extra L-Argwas added.

Method #5 (same as #4 without Na-Bis pre-heating): LD and the respectiveamount of L-Arg were weighed; water was added. The suspension was heatedfor 7 min at 75° C. until fully dissolved followed by 7 min at RT. CDand the respective amount of L-Arg were weighed, and added to the LD-argsolution at 60° C. until fully dissolved. Finally, extra L-Arg wasadded.

After cooling down, all formulations from all methods were divided in to3 vials, and water, Na-Bis solution or Na-Bis-Arg solution was added toeach vial. The physical and chemical stability were evaluated and arepresented in Table A1 and A2:

TABLE A1 Physical stability: First test Stability Second test Stability24 48 72 24 48 Method hours hours hours hours hours 1 Water +++ NR NR ++NR Na-Bis solution +++ ++ Na-Bis solution +++ ++ titrated with L-Arg 2Water + ++ NR − +/− Na-Bis solution − + − +/− Na-Bis solution − +/− −+/− titrated with L-Arg 3 Water − − + − Very few Na-Bis solution − − +(more − particles than at the 13, 15) bottom Na-Bis solution − − + −titrated with L-Arg 4 Water + NR NR + NR Na-Bis solution + + Na-Bissolution +/− + titrated with L-Arg 5 Water ++ NR NR + NR Na-Bis solution++ + Na-Bis solution ++ + titrated with L-Arg − No precipitate +Precipitate

The formulations were sampled for HPLC analysis at the end of thepreparation and after 5 days of stability at RT. The recovery after 5days at RT was calculated compared to T=0.

TABLE A2 Chemical Stability First test Second test LD CD LD CD recoveryrecovery recovery recovery after 5 after 5 after 5 after 5 Method days(%) days (%) days (%) days (%) 1 Water 90.6 98.0 89.5 100.4 Na-Bissolution 90.6 98.6 87.0 101.3 Na-Bis solution 90.8 98.0 88.9 99.9titrated with L-Arg 2 Water 98.4 98.2 99.1 100.1 Na-Bis solution 98.298.1 99.4 100.5 Na-Bis solution 99.0 98.5 98.9 99.5 titrated with L-Arg3 Water 99.7 97.5   95.5[1] 96.5 Na-Bis solution 99.2 97.7  97.7^((a))99.1 Na-Bis solution 99.5 98.1  94.9^((a)) 96.2 titrated with L-Arg 4Water 97.7 97.5 96.3 99.3 Na-Bis solution 96.0 95.8 94.9 97.6 Na-Bissolution 97.7 97.9 96.3 100.0 titrated with L-Arg 5 Water 97.9 96.3 98.1100.9 Na-Bis solution 98.2 98.0 98.2 102.2 Na-Bis solution 97.4 96.798.3 100.6 titrated with L-Arg [1]The recovery values were lower at thesecond test compared to the first test, due to technical problem whichoccurred during the sampling. ^((a))The recovery values were lower atthe second test compared to the first test, due to technical problemwhich occurred during the sampling.

The results in Table A1 and A2 clearly show that the method offormulation preparation has a significant impact on its physical andchemical stability. The formulation of Method #3 shows significantlymore stability.

Example 3: Effect of Arginine on Long Term Stability of Levodopa andLevodopa/Carbidopa Compositions

Liquid formulations with levodopa, carbidopa and arginine were preparedusing the procedure outlined in Example 2, and comparative studies onformulations with a different concentration of arginine and/or an aminosugar (e.g., meglumine), and/or a sugar (e.g. dextrose), and/or a base(NaOH), or another basic amino acid (e.g. lysine, histidine) wereprepared. The results are shown in Table B.

Table B indicate that arginine forms stable solutions with highconcentrations of levodopa and carbidopa (>2.5%) at molar ratios <1:2.5,whereas with other basic amino acids LD does not even dissolve underthese conditions. At molar ratios of LD/CD to arginine 1: <2, thesolutions do not have long term stability, unless meglumine or anothercounterion is used, and meglumine may be used to reduce the molar ratioof arginine to LD/CD.

TABLE B Amino Acid (AA) Other LD/CD Molar Molar Physical Conc. Conc.ratio Conc. ratio stability (%) Name (%) (API:Arg) Name (%) (API:CI)Dissolution at RT 10/0  Lys 8.5 1:2.5 — — — No NA 5/0 Lys 9.25 1:2.5 — —— No NA 3.3/0  Lys 6.2 1:2.5 — — — No NA 3/0 Lys 5.6 1:2.5 — — — PartialNA 2.5/0  Lys 4.6 1:2.5 — — — Yes 2 days 5/0 His 9.8 1:2.5 — — — No NA2.5/0  His 4.9 1:2.5 — — — No NA 1.25/0   His 2.5 1:2.5 — — — Yes 14days 9/0 Arg 8.2 1:1  — — — No NA 4.7/0  Arg 4.0 1:1  — — — No NA 9.5/0 Arg 15.9 1:1.9 — — — Yes 2 days 4.8/1.4 Arg 11.0 1:2.0 — — — Yes ≥2months 4.8/1.4 Arg 12.1 1:2.2 — — — Yes ≥2 months 4.8/1.4 Arg 12.7 1:2.4— — — Yes ≥2 months 5.4/1.5 Arg 13.5   2.1 — — — Yes ≥2 months 5.4/1.5Arg 14.8   2.3 — — — Yes ≥2 months  6/1.5 Arg 14.8   2.1 — — — Yes ≥1month  6/1.5 Arg 16.0   2.3 — — — Yes ≥2 months 7/2 Arg 17.8   2.2 — — —Yes ≥1 month  7/1.5 Arg 14.1 1:1.8 Dex 5.0 — Yes Color change  8/1.5 Arg15.7 1:1.9 Dex 5.0 — Yes Color change  10/1.5 Arg 19.2 1:1.9 Dex 5.0 —Yes Color change  6/1.5 Arg 9.3 1:1.5 NaOH 4.6 1:0.5 Yes ≥3 months 5/0 —— — Meg 5.0 1:1  No NA 5/0 — — — Meg 5.9 1:1.2 No NA 5/0 — — — Meg 10.8 1:2.2 Yes NA  8/1.5 Arg 15.7 1:1.9 Meg 3.2 1:0.4 Yes ≥4.5 months  8/1.5Arg 12.2 1:1.5 Meg 7.9 1:1  Yes ≥4.5 months  10/1.5 Arg 19.2 1:1.9 Meg4.0 1:0.4 Yes ≥4.5 months  10/1.5 Arg 14.6 1:1.5 Meg 9.9 1:1  Yes ≥4.5months  7/1.5 Arg 14.1 1:1.9 Meg 2.8 1:0.4 Yes ≥4.5 months  7/1.5 Arg10.7 1:1.5 Meg 6.9 1:1  Yes ≥4.5 months Lys—Lysine; His—Histidine;Arg—Arginine; Dex—Dextrose; Meg—Meglumine.

Liquid formulations were prepared by weighing all powders (LD, CD andL-Arg) and the addition of water pre-heated to 73±3° C. Suspension wasput in a water bath at 73±3° C. and stirred for 10 min until fullydissolved. LD/CD solution was kept at RT for 10 min to cool down. Then,ascorbic acid was added. Solutions were divided in to glass vials andkept at +25° C. and at −20° C. for the indicated period of time. Priorto analyses, frozen vials were placed at RT until fully thawed.Formulations were then mixed and subjected to stability analyses.

Tables C indicate the effect of 1-arginine on physical and chemical longterm stability at +25° C. and at −20° C.

Liquid formulations were prepared by weighing all powders (LD, CD andL-Arg) and the addition of water pre-heated to 73±3° C. Suspension wasput in a water bath at 73±3° C. and stirred for 10 min until fullydissolved. LD/CD solution was kept at RT for 10 min to cool down. Then,ascorbic acid was added. Solutions were divided in to glass vials andkept at +25° C. and at −20° C. for the indicated period of time. Priorto analyses, frozen vials were placed at RT until fully thawed.Formulations were then mixed and subjected to stability analyses.

TABLE C1 L-Arg Physical Stability (% from T = 0) at RT conc. stability 5days 2 months Formulation (%) at RT LD CD LD CD 6/1.5% LD/CD 13.5 6 days100.0 97.5 (1% Na-Asc) 14.2 At least 100.8 96.7 14.8 7 days 99.6 96.616.0 99.5 96.6 4.8/1.4% LD/CD 11.0 At least 99.4 97.3 100.1 93.7 (1%Na-Asc) 11.6 2 months 98.9 97.4 100.6 96.2 12.1 99.1 97.0 100.3 94.312.7 99.4 97.2 99.0 92.4

TABLE C2 Stability (% from T = 0) 2 weeks at −20 ± 5° C. L-ArgImmediately 24 hours conc. Physical after thawing at RT Formulation (%)stability LD CD LD CD 6/1.5% LD/CD 13.5 At least 24 99.7 98.4 100.0 99.1(1% Na-Asc) 14.2 hr after 99.8 98.1 101.0 99.4 at −20° C. 14.8 thawing100.0 98.9 99.9 98.9 16.0 99.9 98.8 100.3 99.3

TABLE C3 L-Arg Physical stability (at RT) Formulation conc. (%) 1%Na-Asc 1% Asc 6/1.5% LD/CD 14.8 At least 3 weeks At least 3 days 15.816.8 5.4/1.5% LD/CD 12.3 At least 3 days 13.5 14.8

TABLE C4 Physical L-Arg stability Stability (% from T = 0) at RT conc.(after 2 1 weeks 2 weeks 1 month Formulation (%) month at RT) LD CD LDCD LD CD 5.4/1.5% LD/CD 13.5 + 101.4 100.4 101.7 98.4 98.8 103.1 (1%Asc) 14.8 + 101.4 101.4 102.0 100.1 99.0 104.2 6/1.5% LD/CD 14.8 + 101.8101.5 101.6 99.6 99.0 104.2 (1% Asc) 16.0 − 101.1 100.4 102.8 100.6 99.4104.2 7/2% LD/CD 17.8 + 101.7 101.0 102.7 99.7 98.7 103.1 (1% Asc) 7/2%LD/CD − 100.6 NA 101.9 99.2 98.4 103.6 (1% Na-Asc)

TABLE C5 Physical Stability (% from T = 0) Stability (% from T = 0)L-Arg Stability 2 weeks at −20 ± 5° C. 5 weeks at −20 ± 5° C. conc. (11days after immediately after thawing immediately after thawingFormulation (%) thawing) LD CD LD CD 5.4/1.5% LD/CD 13.5 + 102.3 99.599.4 104.3 (1% Asc) 14.8 − 102.7 101.3 99.6 104.6 6/1.5% LD/CD 14.8 −102.6 101.1 99.1 104.2 (1% Asc) 16.0 − 103.2 100.9 99.2 104.3 7/2% LD/CD17.8 + 102.8 101.0 99.2 104.3 (1% Asc) 7/2% LD/CD − 102.9 101.0 99.4104.4 (1% Na-Asc)

TABLE C6 LD/CD conc. L-Arg conc. (%) Physical Stability at 25° C. 12/3%24.4 Considerable precipitate on Day 5 29.6 Slight precipitate on Day 532.1 No precipitate on Day 7

Tables C1-C6 indicate that there is a correlation between the molarratio of arginine to LD/CD and stability where generally compositionshaving more arginine, have longer stability: LD/CD:arginine solutions(at molar ratios of 1:≥2.1) are stable for at least 1 month at RT and at−20±5° C. The solutions are stable even at very high solidconcentrations (total of >45%).

Formulations containing 6/1.5% and 5.4/1.5% LD/CD and varying L-Argconcentrations were titrated with Acetic acid (100%) or Lactic acid(85%) to investigate the effect of pH and L-arginine concentration onthe physical stability of the solutions. Table D indicates the results.

TABLE D pH L-Arginine Asc/ pH Lactic after pH 4 24 (%) Na-Asc before (%)Lactic drop hours hours 6/1.5% 14.8 Na-Asc 9.53 1.1 9.25 −0.28 OK +LD/CD 9.53 1.7 9.16 −0.37 + + 9.53 2.3 9.02 −0.51 ++ + 14.8 Asc 9.410.85 9.24 −0.17 OK + 9.42 1.3 9.14 −0.28 + + 9.41 1.7 9.06 −0.35 + +15.8 Na-Asc 9.52 1.1 9.33 −0.19 OK OK 9.50 1.7 9.21 −0.32 OK + 9.53 2.39.08 −0.45 + + 15.8 Asc 9.44 0.85 9.27 −0.17 OK OK 9.45 1.3 9.19 −0.26OK + 9.45 1.7 9.11 −0.34 + + 16.8 Na-Asc 9.56 1.1 9.36 −0.20 OK OK 9.561.7 9.23 −0.33 OK OK 9.56 2.3 9.09 −0.47 OK + 16.8 Asc 9.46 0.85 9.30−0.16 OK OK 9.46 1.3 9.20 −0.26 OK OK 9.47 1.7 9.11 −0.36 OK +L-Arginine Asc/ pH Lactic Acetic pH pH 2 3 10 (%) Na-Asc before (%) (%)after drop days days days 5.4/1.5% 12.3 Na-Asc 9.41 0.36 — 9.35 −0.06OK + + LD/CD 9.43 1.0  — 9.18 −0.25 ++ + + 9.43 — 0.35 9.29 −0.14 OK + +12.3 Asc 9.28 0.36 — 9.20 −0.08 ++ + + 9.29 1.0  — 9.05 −0.24 ++ ++ ++9.29 — 0.35 9.14 −0.15 ++ ++ ++ 13.5 Na-Asc 9.50 0.36 — 9.38 −0.12 OK OKOK 9.48 1.0  — 9.25 −0.23 + + + 9.49 — 0.35 9.35 −0.14 OK OK OK 13.5 Asc9.32 0.36 — 9.25 −0.07 + + + 9.33 1.0  — 9.11 −0.22 ++ ++ ++ 9.34 — 0.359.20 −0.14 + + + 14.8 Na-Asc 9.51 0.36 — 9.43 −0.08 OK OK OK 9.51 1.0  —9.28 −0.23 OK OK OK 9.51 — 0.35 9.38 −0.13 OK OK OK 14.8 Asc 9.36 0.36 —9.29 −0.07 OK OK OK 9.37 1.0  — 9.13 −0.24 +/− + + 9.36 — 0.35 9.23−0.13 OK OK OK OK—no precipitate; +/− very few particles; + slightprecipitate; ++ considerable precipitate

Table E shows the physical and chemical stability 3 weekspost-preparation of the 6/1.5/14.8% LD/CD/Arg formulation used for thestability tests shown in Table D.

TABLE E Physical Stability Asc/Na-Asc stability (% of T = 0) Formulation(1%) (at RT) LD CD 6/1.5% LD/CD, Asc ≥3 weeks 103.1 98.9 14.8% L-ArgNa-Asc 101.1 97.4

Table D indicate that ascorbic acid reduces the pH by 0.1-0.15 units ascompared to Na-ascorbate and that other organic acids can further reducethe pH of the formulations. But the physical stability test resultsindicate that formulations are not generally stable at pH <9.15±0.5.Formulations with Na-ascorbate appear more stable than formulations withascorbic acid at a given L-arginine concentration. Thus, it is suggestedthat excess of acid may cause precipitation in the absence of adequateamount of L-Arg.

Example 4: Stability of Levodopa Formulations with Carbidopa In-Vitroand Ex-Vivo

The effect of carbidopa on levodopa formulations was investigated.Levodopa (LD) formulations were prepared with 0, 0.5, 1, 1.5 & 2% byweight carbidopa (CD) and a constant concentration of arginine. Physicaland chemical stabilities were evaluated, as shown in Table F:

TABLE F Stability (% from T = 0) Physical 3 days 15 days FormulationN₂+/− stability LD CD LD CD 7% LD w/o CD + Stable 99.2 NA 103.4 NA −Stable 98.1 NA — NA 0.5% CD + Stable 98.6 94.7 104.1 108.1 − Stable 98.795.6 — — 1% CD + Stable 98.9 95.2 102.5 104.4 − Slight 97.9 94.0 — —precipitate 1.5% CD + 7 days 98.1 94.2 103.7 104.8 − 99.6 96.0 — — 2%CD + 4 days 98.9 94.5 102.9 103.3 − 98.3 94.8 — —

The experimental results shown in FIG. 1A (see figures) indicate thatcarbidopa prevented dark yellow color formation in the presence of air,in a dose related manner. In the absence of air (with N₂ in the headspace) 0.5% CD was sufficient to inhibit this color formation. It issuggested that CD inhibits oxidation of LD in vitro. The experimentalresults shown in Table F indicate that carbidopa does not have asignificant effect on the chemical stability of levodopa. It also showsthat the ratio between arginine and the total active ingredients isimportant to prevent precipitation, i.e., the physical stability of theformulation depended on the relative concentration of arginine

In an additional experiment, LD formulations were prepared with 0, 0.5,1 & 2% CD and respective concentrations of arginine. Physical andchemical stability were evaluated, and results are shown in Table G:

TABLE G Physical Chemical Stability at RT (% of t₀) stability at RTL-Arg 3 days 1 month after thawing 1 month after Formulation (%) LD CDLD CD thawing LD 6% LD/0% CD 13.5 102.3 — 6% LD/0% CD 13.5 102.3 6%LD/0.5% CD 14.2 103.3 100.4   6% LD/0.5% CD 14.2 103.3 6% LD/1% CD 14.8103.5 101.3 6% LD/1% CD 14.8 103.5 6% LD/2% CD 16.5 103.3 101.6 6% LD/2%CD 16.5 103.3

In the presence of adequate concentrations of L-arginine, allformulations ex-vivo were stable for at least a month at RT followingthawing, as shown in Table G.

The effect of carbidopa on the stability of levodopa formulations isshown in FIG. 1B. A 7% LD-arginine solution, with or without 2% CD, wascontinuously administered at 0.08 ml/h×18 h, 37° C. into a 5×5 cm fresh,full-thickness pig skin. The right hand side of FIG. 1B indicates thelack of black by-products formation, suggesting that CD inhibitsoxidation of LD ex vivo and may also inhibit the formation of o-quinonesand melanin.

Example 5: Stability of Carbidopa Formulations with Levodopa

The effect of levodopa on the stability of carbidopa was investigated.Table H indicates results.

TABLE H T = 0 T = 4 days at 25° C. LD CD LD CD Recovery of CDFormulation (mg/g) (mg/g) (mg/g) (mg/g) (% of t0) 6% LD/2% CD 60.3 19.4Air 63.2 18.9 97.4 N₂ 62.9 19.0 97.9 2% CD N/A 19.5 Air N/A 15.9 81.5 N₂N/A 19.0 97.4 T = 0 Retention time (srea of impurity) Formulation 3.383.54 4.2 4.85 5.2 5.52 5.77 12.10 13.35 13.60 14.60 6% LD/2% CD NA NA1.08 3.15 1.67 0.34 0.86 NA 1.48 0.95 1.63 2% CD 1.30 0.25 NA 1.79 NA NA0.95 0.35 NA 1.45 3.83 CD vs. CD/LD 0.6 1.1 1.5 2.3 T = 4 days at 25° C.Retention Time (Area of Impurity) 3.15 3.32 4.12 4.82 5.65 11.92 12.1012.27 12.70 13.53 14.55 6% LD/2% CD Air 12.23 1.00 2.10 3.57 1.94 0.790.69 0.89 1.34 1.34 16.82 N₂  8.09 0.82 1.48 3.63 1.61 0.44 0.53 0.560.56 1.08 11.82 2% CD Air NA 1.59 NA 9.49 1.18 NA NA NA 7.54 24.04 70.22N₂ NA 1.65 NA 6.63 1.07 0.23 NA NA 0.50 3.62 25.45 CD vs. CD/LD Air 1.62.7 0.6 5.6 17.9 4.2 N₂ 2.0 1.8 0.7 0.5  0.9 3.4 2.2

Table H indicates that CD was less sensitive to oxidation anddegradation and was more stable in the presence of LD: The area ofimpurities at R.T. 4.82, 5.65, 12.7, 13.53 and 14.55 were significantlyincreased under aerobic conditions when LD was not present, and the areaof impurities at R.T. at 4.82 and 13.53 were increased even in theabsence of oxygen. It appears that LD may protect CD from degradation.

Example 6: Toxicity and Pharmocokinetics of Levodopa Formulations withCarbidopa

The effect of carbidopa on levodopa local toxicity was investigated inpigs: Solutions containing 6% LD and 0, 0.5 or 1% CD with the respectiveamount of arginine (13.5, 14.2 or 14.8%, respectively) were continuouslyadministered SC to pigs at 0.16 ml/h×24 h. Each formulation wasadministered to 2 pigs. Skin samples were collected 8±1 days thereafter.FIG. 2 shows that the presence of 1% carbidopa reduces the severity andextent of levodopa dependent toxicity, in-vivo.

The effect of carbidopa on the pharmacokinetics of levodopa andcarbidopa were investigated. Solutions containing 6% LD and 0, 0.5, 1 or2% CD and the respective amount of arginine (13.5, 14.2, 14.8 or 16.5%respectively) were continuously administered SC to pigs at 0.16 ml/h×24h. FIGS. 3A-3C show that CD has a significant effect on thepharmacokinetics of LD. This effect was dose dependent and linearbetween ±0.3 and ±1.2% CD, as in example 6.

Example 7: Effect of Tyrosinase Inhibitors

The effect of tyrosinase inhibitors, substrate analogues, Cu′ chelatorsand 0-quinone scavengers on oxidation on levodopa (LD) followingcontinuous subcutaneous administration of levodopa/carbidopaformulations at 37° C. in to the subcutaneous tissue of pig skin, wasstudied ex-vivo.

Full thickness pig skin samples (including the subcutaneous tissue) wereplaced on top of a 100 ml glass bottle filled to the top with warm PBS,in an incubator set at 37° C. The skin was directly in contact with thePBS, and the skin and bottle were then covered with parafilm to protectthe inner side of the skin from air as much as possible. Theformulations were administered subcutaneously using a 22 G butterfly andan infusion pump set at 0.08 ml/h.

A list of various potential levodopa stabilizers andoxidation/degradation/metabolism inhibitors are listed in Table I. Theeffect of representatives from each group and combinations there offwere tested ex vivo and is shown in FIGS. 4A-4D.

The number of each skin sample corresponds to the numbered formulationthat was administered as per Table J below:

TABLE I Cu⁺⁺ chelators Clinical use Chemical Min. Max. pH name MWSolubility dose dose compatibility EDTA calcium 374 Soluble 500 mg/m20.17%  √ disodium EDTA 372 Soluble NA 0.2% √ disodium DMSA (succimer)182 Soluble 10 mg/kg × 3 NI pKa 3, 3.9 DPA (D- 149 Soluble 125 mg × 1 NApKa 8 & 10.5 penicillamine) Trientine ? Soluble 250 mg × 2 NA (HCl)Dimercaprol 124 0.275% 2.5 mg × 4 NA pKa 10.4 Clioquinol 305 <0.1% 125mg × 2 NA Sodium NA 0.2% thiosulfate TETA NA NA TEPA NA NA Curcumin NANA Neocuproine NA NA Tannin NA NA Cuprizone NA NA Substrate analoguesChemical Name MW Solubility pH compatibility Sodium benzoate Pka 4.2L-phenylalanine Tyrosinase inhibitors Min required Generic name MWSolubility concentration Captopril 217 Soluble 0.04 mg/ml Methimazole114 Soluble Quercetin 302 Arbutin Aloesin N-acetylglucoseamine RetinoicAcid a-tocopheryl ferulate MAP(Mg ascorbyl phosph) O-quinone scavengersClinical use pH Generic name MW Solubility Dose compatibility L-Cysteine121 Soluble 0.1% pKa 8, 10 Ascorbic acid 176 Soluble 1.0% pKa 4, 11Gluthatione (GSH) 0.5% pKa 8, 10 NI—Not indicated; NA—Not applicable

TABLE J # Composition 1 7% LD, 0.02% Na-bisulfite 2 7% LD, 2% CD 3 7%LD, 1% Na-ascorbate 4 7% LD, 0.1% Cysteine 5 7% LD, 0.2% Na₂ EDTA 6 7%LD, 0.2% EDTA-Ca—Na₂ 7 7% LD, 0.2% methimazole 8 7% LD, 0.2%D-penicillamine 9 7% LD, 0.2% Captopril 10 7% LD, 0.2% EDTA-Ca—Na2, 0.2%captopril, 1% Na-ascorbate 11 5.4% LD, 0.02% Na-bisulfite 12 5.4% LD,1.5% CD 13 5.4% LD, 1.5% CD, 1% Na-ascorbate 14 5.4% LD, 1.5% CD, 1%Na-ascorbate, 0.2% EDTA-Ca—Na₂ 15 5.4% LD, 1.5% CD, 1% Na-ascorbate,0.2% captopril 16 5.4% LD, 1.5% CD, 1% Na-ascorbate, 0.2% methimazole 175.4% LD, 1.5% CD, 0.2% captopril 18 5.4% LD, 1.5% CD, 0.2% captopril,0.2% EDTA-Ca—Na₂ 19 5.4% LD, 1.5% CD, 0.2% methimazole 20 5.4% LD, 1.5%CD, 0.2% methimazole, 0.2% EDTA-Ca—Na₂ 21 7% LD, 0.02% Na-bisulfite 227% LD, 2% CD 23 7% LD, 2% CD, 0.2% Ascorbate 24 7% LD, 2% CD, 1%Ascorbate 25 7% LD, 2% CD, 0.2% ascorbate, 0.2% captopril 26 7% LD, 2%CD, 1% Ascorbate, 0.2% Captopril 27 7% LD, 2% CD, 0.2% Ascorbate, 0.2%Na2-EDTA 28 5.4% LD, 1.5% CD 29 5.4% LD, 1.5% CD, 0.2% Ascorbate 30 5.4%LD, 1.5% CD, 1% Ascorbate 31 7% LD, 0.02% Na-bisulfite 32 7% LD, 2% CD33 7% LD, 2% CD, 0.5% ascorbate 34 7% LD, 2% CD, 0.5% ascorbate, 0.2%captopril 35 7% LD, 2% CD, 1% ascorbate 36 7% LD, 2% CD, 1% ascorbate,0.2% captopril 37 7% LD, 2% CD, 1% ascorbate, 0.2% captopril, 0.2%Na₂EDTA 38 7% LD, 2% CD, 1% ascorbate, 0 2% captopril, 0.2% Ca—Na₂EDTA

FIGS. 4A-4C indicate that ascorbate, at a concentration of ≥0.5%, wassufficient to inhibit color change of levodopa and carbidopa in pig skinsamples. Other compounds tested were less effective.

Example 8 Effect of Tyrosinase Inhibitors on Subcutaneous ToxicityIn-Vivo

The effect of tyrosinase inhibitors on subcutaneous toxicity following24 h-continuous subcutaneous administration of LD/CD in Pigs was studiedfor 6 to 11 days after administration. Results are shown in Table K:

TABLE K Histological Score Formulation Inflammation Necrosis LD/CD(5.4/1.5%) + 1% ascorbate 2.2 2.0 LD/CD (5.4/1.5%) + 1% ascorbate, 2.62.5 0.2% Ca—Na₂-EDTA LD/CD (5.4/1.5%) + 1% ascorbate, 2.5 2.6 0.2%Ca—Na₂-EDTA, 0.2% captopril Score Key 0 No lesions 1 Minimalinflammation/necrosis 2 Mild inflammation/necrosis 3 Moderateinflammation/necrosis 4 Severe inflammation/necrosis

Under the experimental conditions employed, captopril and/or Ca—Na₂EDTAdid not have a supplementary effect, as compared to 1% ascorbate alone,in the protection from local toxicity.

Example 9: Plasma Levels of Levodopa Following SubcutaneousAdministration

In this experiment, the purpose was to determine the plasma levels of LD(levodopa) following continuous subcutaneous administration ofcarbidopa, levodopa or entacapone and combinations thereof with oralLD/CD in pigs.

Landrace×Large White female pigs weighing about 22 kg were treated,starting on Day 1 at 15:00 as per table 1, with oral LD/CD 100/25 andwith the respective test formulations, containing carbidopa, levodopa orentacapone and combinations thereof, formulated with arginine, asdescribed above, and administered continuously subcutaneously via adermal patch (Omnipod®) at a rate of 0.08 ml/h.

Table L indicates the treatment protocol of each group. The formulationswere prepared as in Example 1 and 2.

TABLE L Treatment group None CD CD + E E LD + CD LD n 3 3 3 2 2 1 SCroute of No SC 2% carbidopa 2% carbidopa + 10% entacapone 7% levodopa +7% levodopa administration treatment 10% entacapone 2% carbidopa Oraltreatment 100/25 levodopa/carbidopa

Blood samples were collected following the 3^(rd) oral dose atpre-determined time points and plasma levels of levodopa, carbidopa and3-OMD were analyzed by HPLC-ECD.

FIGS. 5A and 5B indicate the mean levodopa plasma concentrationsfollowing oral administration of Sinemet (oral 100/25 LD/CD) withcontinuous SC administration of FIG. 5A) Entacapone (200 mg/24 h)±CD (40mg/24 h) or FIG. 5B) Levodopa (140 mg/24 h)±CD (40 mg/24 h) in pigs (allsubcutaneous formulations included arginine, as above).

Results show that there is a synergistic effect between entacapone (200mg/24 h) and CD (40 mg/24 h) on the plasma PK of levodopa (ng/ml) whenco-administered continuously subcutaneously, as compared to thecalculated LD plasma PK obtained after adding the plasma concentrationsof LD following the continuous SC administration of CD and entacaponeeach alone (FIG. 1A and Table 2, C vs. B+D). Results also show thatthere is an additive effect between levodopa (140 mg/24 h) and CD (40mg/24 h) on the plasma PK of levodopa (ng/ml) when co-administeredcontinuously subcutaneously, as compared to the calculated LD plasma PKobtained after adding the plasma concentrations of LD following thecontinuous SC administration of CD and LD each alone (FIG. 1B and Table2, E vs. D+F). Moreover, the results suggest that continuous SCadministration of LD and CD may be sufficient to maintain constant,continuous levodopa plasma concentrations even in the absence of oralLD/CD administration (FIG. 5B dotted line and Table M ‘E minus A’).Table M presents trough concentrations of plasma levodopa 6½ and 8 hPost-Oral LD/CD administration.

TABLE M SC treatment (LD + CD)-None LD + CD E + CD Time None E E + CD CDLD + CD calculated LD calculated calculated point (h) A B C D E E − A FD + F B + D 6.5 51 179 1695 998 1226 1174 322 1320 1177 8 0 0 1474 8681227 1227 413 1281 868 E—entacapone; CD—carbidopa; LD—levodopa; NA—notAvailable

FIG. 6 shows tissue biopsies from the application site of thelevodopa-carbidopa arginine combination formulation and thelevodopa/arginine formulation. No visible tissue irritation or damagewas apparent in the levodopa-carbidopa arginine formulation. The siteadministered with levodopa-arginine formulation appears to have someblackening of tissue. Without being limited by any theory, it is thoughtthat having carbidopa and arginine together with levodopa (arginine)formulation protects the local tissue from local damage of levodopa bypreventing oxidation of levodopa into irritant by products, and thatcarbidopa is a potent anti-oxidant.

Equivalents

While specific embodiments of the subject invention have been discussed,the above specification is illustrative and not restrictive. Manyvariations of the invention will become apparent to those skilled in theart upon review of this specification. The full scope of the inventionshould be determined by reference to the claims, along with their fullscope of equivalents, and the specification, along with such variations.

Unless otherwise indicated, all numbers expressing quantities ofingredients, reaction conditions, and so forth used in the specificationand claims are to be understood as being modified in all instances bythe term “about.” Accordingly, unless indicated to the contrary, thenumerical parameters set forth in this specification and attached claimsare approximations that may vary depending upon the desired propertiessought to be obtained by the present invention.

INCORPORATION BY REFERENCE

The entire contents of all patents, published patent applications,websites, and other references cited herein are hereby expresslyincorporated herein in their entireties by reference.

1. A pharmaceutically acceptable composition having a pH of about 9.1 toabout 9.8 at 25° C., comprising: active components comprising carbidopaand at least about 4% by weight levodopa; arginine and optionallymeglumine.
 2. The pharmaceutically acceptable composition of claim 1,wherein the molar ratio of active components to the arginine is about1:1.8 to about 1:3.
 3. The pharmaceutically acceptable composition ofclaim 1, comprising about 4% to about 12% by weight levodopa.
 4. Thepharmaceutically acceptable composition of claim 1, comprising about 1%to about 6% by weight carbidopa.
 5. The pharmaceutically acceptablecomposition of claim 1, comprising meglumine.
 6. The pharmaceuticallyacceptable composition of claim 5, wherein the molar ratio of activecomponents to the arginine is about 1:1.1 to about 1:1.9, and the molarratio of active components to the meglumine is about 1:0.3 to about1:1.5.
 7. The pharmaceutically acceptable composition of claim 5,wherein the molar ratio of active components to the meglumine is about1:0.3 to about 1:1.2.
 8. The pharmaceutically acceptable composition ofclaim 5, wherein the composition comprises about 2.0% to about 11% byweight meglumine.
 9. The pharmaceutically acceptable composition ofclaim 1, comprising about 10% to about 35% by weight arginine.
 10. Thepharmaceutically acceptable composition of claim 1, further comprisingan agent that inhibits the formation of oxidation products.
 11. Thepharmaceutically acceptable composition of claim 10, wherein the agentis selected from ascorbic acid, Na-ascorbate, L-cysteine,N-acetylcysteine (NAC), glutathione (GSH), Na₂-EDTA, Na₂-EDTA-Ca, orcombinations thereof.
 12. The pharmaceutically acceptable composition ofclaim 10, wherein said agent is ascorbic acid or a pharmaceuticallyacceptable salt thereof.
 13. The pharmaceutically acceptable compositionof claim 1, further comprising sodium bisulfite.
 14. A pharmaceuticallyacceptable composition comprising levodopa, arginine and optionallymeglumine; and further comprising ascorbic acid or a pharmaceuticallyacceptable salt thereof.
 15. The pharmaceutically acceptable compositionof claim 14, wherein the ascorbic acid salt is selected from ascorbate,sodium ascorbate, calcium ascorbate, potassium ascorbate, ascorbylpalmitate, or ascorbyl stearate. 16-25. (canceled)
 26. A transdermalpatch suitable for administering a pharmaceutically acceptablecomposition according to claim
 1. 27. A method of treating aneurological or movement disorder in a patient in need thereof, themethod comprising the steps of: (a) substantially continuouslyadministering to said patient a composition of claim 1, wherein thecomposition is administered subcutaneously, intraduodenally orintravenously; and (b) orally administering levodopa and/or carbidopaand optionally entacapone or tolcapone. 28-32. (canceled)
 33. The methodof claim 27, wherein the rate of administering the composition is about1.0±0.5 ml/hour during the day or during patient activity, and about 0to about 0.5 ml/hour at night or at rest.
 34. The method of claim 27,wherein the disorder is Parkinson's disease.
 35. A pharmaceuticallyacceptable formulation comprising about 2.5 to about 7% by weightlevodopa, about 0 to about 2% by weight carbidopa, about 5 to about 18%by weight arginine, and about 0.25% to about 3% by weight ascorbic acidor a pharmaceutically acceptable salt thereof. 36-47. (canceled)